We present in situ studies on the self-assembly and,dynamic evolution of collagen gels from semidilute solutions in a microfluidic device. Collagen fibrils not only reinforce the mechanical properties of bone and tissues, but they also influence cellular motility and morphology. We access the initial steps of the hierarchical self-assembly of collagen fibrils and networks by using hydrodynamic focusing to form oriented fibers. The accurate description of the conditions within the microchannel requires a numerical expression for the pH in the device as well as a modified mathematical description of the viscosity, which increases nearly 300-fold as collagen fibrils form around neutral pH. Finite element modeling profiles overlay impressively with cross-polarized microscopy images of the birefringent fibrils in the channel. Real-time X-ray microdiffraction measurements in flow indicate an enhanced supramolecular packing having a unit spacing commensurate with that of a pentameric collagen subunit. These results have significant implications for the field of biomedicine, wherein new aligned, cellularly active, and mechanically strengthened materials continue to be in demand. However, this work is also-remarkable from a more fundamental, biophysical point of view because the underlying concepts may be generalized to a large pool of systems.